Developmental exposure to natural or environmental estrogens predisposes to prostate carcinogenesis with aging; however, the molecular underpinnings of this phenomenon are unclear. We present evidence that developmental reprogramming of the prostate by estrogens may be mediated, in part, through epigenetic alterations. Using methylation-sensitive fingerprinting (MSRP) as an initial screen for genome-wide methylation changes, we identified multiple prostatic genes whose methylation status was permanently altered in rats as a result of neonatal estradiol and bisphenol A (BPA) exposures at environmentally relevant doses. Detailed characterization of phosphodiesterase 4D4 (PDE4D4), an enzyme involved in cAMP breakdown, revealed aberrant promoter CpG island methylation patterns with resultant changes in gene transcription as the animals aged. Importantly, these epigenetic alterations were associated with increased susceptibility to hormonal carcinogenesis of the rat prostate gland. Thus we hypothesize that hormonal and environmental factors during the sensitive developmental period permanently impact the prostate epigenome which, in turn, promotes prostate disease with aging. The objectives of the present proposal are to characterize in detail the prostatic gene methylation and transcriptional alterations which result from early - life estrogenic exposures (natural and environmental) and to determine if there is an epigenetic basis for estrogenic predisposition to carcinogenesis. Three Aims are proposed: Aim 1: Determine and characterize gene methylation targets in the rat prostate gland which are sensitive to neonatal estrogen or BPA exposure and result in altered gene expression. Aim 2: Directly assess whether epigenetic regulation of identified candidate genes results in increased carcinogenic potential in the prostate gland. Aim 3: Determine if epigenomic alterations in the prostate by natural or environmental estrogens impact genomic susceptibility to prostate cancer. The general approach is as follows: Outbred and inbred rats will be neonatally exposed to estradiol or BPA and hormonal carcinogenesis will be induced in the adult animal. MSRP will be used to expand our prostatic screen to identify a full panel of candidate genes and a stringent algorithm will be followed to identify candidates with regulatory CpG islands. Site-specific methylation and resultant transcriptional regulation will be confirmed. The carcinogenic roles of the candidates will be elucidated by correlative studies in vivo and transgene in vitro experiments with cell and explant cultures. Identification of methylation fingerprints and specific genes with permanent methylation alterations may serve as molecular markers for developmental estrogenic exposures and provide molecular insight into the epigenomic plasticity that predisposes to prostate cancer with aging. The findings will serve as a model for human exposures to prevalent environmental endocrine disrupters (e.g. BPA) with suspected carcinogenic potential.